Malaria parasites are responsible for 300-500 million infections and 2-3 million deaths annually. Transmission of malaria between vertebrate hosts involves an obligate sexual developmental cycle in the anopheline mosquito vector. The sexual stages are absolutely essential for malaria transmission. The molecular mechanisms underlying sexual differentiation and development in Plasmodium falciparum remain largely unknown. After the initial commitment to the sexual cycle, Plasmodium gametocytes undergo what appears to be a multi-step growth and development process, possibly involving several gene products. The goal of this proposal is to investigate the functional involvement of two proteins (Pfg27 and Pfs16), abundantly expressed early during gametocytogenesis. We have recently disrupted the gene for Pfg27 by homologous recombination, resulting in the loss of sexual phenotype in the transformed parasites. These studies for the first time have shown that Pfg27, a protein expressed in stage I and II gametocytes is critical for gametocyte development. Studies on Pfs16 disruption will further elucidate sexual differentiation process in P. falciparum. Investigations on stably transformed parasites will also offer an opportunity to evaluate transcriptional control mechanisms, sexual stage- specificity of promoters and complementation of disrupted genes. A combination of immunochemical and molecular (yeast-two-hybrid) approaches will be employed to investigate protein-protein interactions involving Pfg27 and Pfs16 and other cellular proteins. These studies could possibly identify novel proteins as interacting functional partners, thus advancing our understanding of the biological roles of Pfg27 and Pfs16 in the sexual differentiation and development of transmission competent parasites. The proposed studies will thus offer a novel molecular genetic approach to dissect mechanisms underlying sexual development in P. falciparum. Such information could lead to rational development of immunological and/or chemotherapeutic arsenal to prevent transmission of P. falciparum malaria.